107 / 328
Primary and Secondary Evaluation of Method SJW-02
SJW-02:
Evaluation of major active components in St. John’s wort dietary supplements by high-
performance liquid chromatography with photodiode array detection and
electrospray mass spectrometric confirmation
Author(s):
Frances F. Liu\ Catharina Y.W. Ang*, Thomas M. Heinze, Joshua D. Rankin2,
Richard D. Beger, James P. Freeman, Jackson O. Lay Jr., US Food and Drug
Administration, National Center for Toxicological Research, Division of Chemistry, 3900
NCTR Road, Jefferson, AR 72079, USA
S
UMMARY OF
M
ETHOD
:
G
ENERAL
C
OMMENTS
:
This method was published in 2000 and used RP-HPLC/DAD for the quantification of the flavonoids
(rutin, hyperoside, isoquercitrin, quercitrin, and quercetin), phloroglucinol (hyperforin), and
naphthodianthrones (hypericin and pseudohypericin) in five capsule products of St. John's wort. LC-ESI
MS was used for confirmation of the detected compounds. Samples are extracted with an ethanol-
acetone solution via shaking in a water bath at 55°C for 5.6 hours. Extract is filtered through filter paper
into volumetric flask and diluted to 50.00 mL prior to analysis by reversed phase HPLC. Chromatographic
separation is achieved on a 3 μm 250 x 4 mm C18 column at 25°C employing a gradient with
0.5%TFA/water and 0.5% TFA in methanol/acetonitrile (13:7). Analytes are detected by photodiode
array at 284 and 590 nm and quantified using luteolin as an internal standard with peak confirmation by
subsequent electrospray ionization mass spectrometry. The LC-MS method is 125 minutes long,
employs a different chromatographic conditions, as opposed to detection in series, and as such offers no
real advantage.
P
ROS
/S
TRENGTHS
:
This method used LC-ESI MS to confirm the identity of the peaks and the purity of the standards was
confirmed by UV, MS and NMR. It detects and quantifies the analytes of interest and has a simple
extraction procedure.
C
ONS
/W
EAKNESSES
:
This method has a long extraction procedure and HPLC run time. Only capsules were used in this
method and the composition of those capsules was not detailed. There were challenges regarding the
precision and stability. The LOQ, RSD
r
, RSD
R
did not appear to meet the SMPR. There was no recovery or
LOD data provided.
EXPERT REVIEW PANEL VOTE
AND
RECOMMENDATION
MOTION: Not to consider this method for First Action Official Method status.
Brown, Schaneberg (Unanimous) Motion Passed
ERP PROFILE SUMMARIES
84